人SP-B蛋白转基因小鼠及细菌性肺炎模型的构建

doi:10.13523/j.cb.20171009

中国生物工程杂志

2017, Vol. 37

Issue (10): 65-71 DOI: 10.13523/j.cb.20171009

技术与方法

人SP-B蛋白转基因小鼠及细菌性肺炎模型的构建

葛林¹, 刘新宇², WANG Guirong³

1. 天津医科大学生物化学与分子生物学系天津 300070;
2. 天津医科大学代谢病医院内分泌研究所卫生部激素与发育重点实验室天津市代谢性疾病重点实验室天津 300070;
3. 美国纽约州立大学上州医科大学纽约 13210

Construction of Human SP-B Protein Transgenic Mice and Bacterial Pneumonia Model

GE Lin¹, LIU Xin-yu², WANG Guirong³

1. Tianjin Medical University, Department of Biochemistry and Molecular Biology, Tianjin 300070, China;
2. Key Laboratory of Hormones and Development Ministry of Health, Tianjin Key Laboratory of Metabolic Diseases, Tianjin Metabolic Diseases Hospital & Tianjin Institute of Endocrinology, Tianjin Medical University, Tianjin 300070, China;
3. SUNY Upstate Medical University, Department of Surgery, Syracuse NY 13210, USA

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摘要： 目的：构建携带人SP-B蛋白+1580 SNP不同等位基因的转基因小鼠并进行细菌性肺炎模型的造模。方法：利用受精卵原核注射技术将hSP-B基因整合至小鼠染色体上获得F₀代小鼠，将其与mSP-B基因敲除鼠进行交配，逐步去除转基因小鼠体内mSP-B基因。利用PCR技术鉴定小鼠基因型，通过测序确定+1580位点的等位基因。将铜绿假单胞菌经支气管灌注接种至小鼠肺内进行细菌性肺炎造模，对照组注射等量灭菌生理盐水。结果：F₂代小鼠只表达人SP-B蛋白而不表达鼠SP-B蛋白，蛋白表达量与人肺内含量相近，即为构建成功的转基因小鼠。3个小鼠家系+1580位点等位基因为T，1个家系为C。细菌接种（1×10⁶ CFU/mouse）后24小时，小鼠肺泡内炎症渗出明显，大量中性粒细胞浸润，SP-B蛋白含量明显降低，但不同等位基因间在此条件下无明显差异。结果：成功构建只表达人SP-B蛋白的转基因小鼠模型，细菌性肺炎模型造模成功，为今后进一步研究人SP-B蛋白的生理功能及+1580基因多态性与肺疾病的关系提供了有力的工具。

关键词： SNP; 细菌性肺炎; SP-B蛋白; 转基因小鼠

Abstract: Objective:to construct the transgenic mice expressing human SP-B gene with different alleles in +1580 SNP and the bacterial pneumonia model. Methods:The hSP-B gene was integrated into the mouse chromosome to obtain the F₀ generation mice by microinjection technology. The mice were mated with mSP-B gene knockout mice to eliminate the mSP-B gene gradually. Using PCR technique to identify the genotype of mice, and to determine the allele of +1580 locus by sequencing. The Pseudomonas aeruginosa was inoculated into the lung of mice to make the model of bacterial pneumonia, and the control group was injected with the same amount of sterile saline. Results:F₂ mice expressed human SP-B protein only, and the expression level of SP-B protein was similar to that in human lung. The +1580 locus alleles T was found in 3 mice families and allele C was found in 1 mice family. After infection for 24 hours, the inflammatory exudation in the alveolar of mice was obvious. A large number of neutrophils were observed in mice alveolus. SP-B protein level was significantly reduced, but there was no significant difference between the different alleles. Conclusions:the construction of human SP-B protein expression transgenic mice model and bacterial pneumonia model were successful. The mice will be a powerful tool for the further studies on physiological function of human SP-B protein and the relationship between +1580 gene polymorphism and lung diseases.

Key words: SP-B protein SNP Transgenic mice Bacterial pneumonia

收稿日期: 2017-04-11 出版日期: 2017-10-25

ZTFLH:

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通讯作者: WANG Guirong,wangg@upstate.edu E-mail: wangg@upstate.edu

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引用本文:

葛林, 刘新宇, WANG Guirong. 人SP-B蛋白转基因小鼠及细菌性肺炎模型的构建[J]. 中国生物工程杂志, 2017, 37(10): 65-71.

GE Lin, LIU Xin-yu, WANG Guirong. Construction of Human SP-B Protein Transgenic Mice and Bacterial Pneumonia Model. China Biotechnology, 2017, 37(10): 65-71.

链接本文:

https://manu60.magtech.com.cn/biotech/CN/10.13523/j.cb.20171009 或 https://manu60.magtech.com.cn/biotech/CN/Y2017/V37/I10/65

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